Renin inhibitors containing alpha-heteroatom amino acids

ABSTRACT

The invention concerns novel renin-inhibitory peptides which contain novel amino acids having an α-heteroatom attached to the backbone of the amino acid. These novel amino acids are positioned at the P 2  position of the peptide. These are useful for treating renin-associated hypertension, hyperaldosteronism, and congestive heart failure. Processes for preparing the peptides, compositions containing them and methods of using them are included. Novel intermediates are also disclosed. Also included is a diagnostic method which uses the compounds to determine the presence of renin-associated hypertension or hyperaldosteronism.

This is a continuation-in-part of United States application Ser. No.154,727 filed Feb. 11, 1988.

BACKGROUND OF THE INVENTION

Renin is a natural enzyme which is released into the blood from thekidney It cleaves its natural substrate, angiotensinogen, releasingdecapeptide, angiotensin I. This is in turn cleaved by converting enzymein the lung, kidney and other tissues to an octapeptide, angiotensin II.Angiotensin II raises blood pressure both directly by causing arteriolarconstriction and indirectly by stimulating release of thesodium-retaining hormone aldosterone from the adrenal gland causing arise in extracellular fluid volume. Inhibitors of renin have been soughtas agents for control of hypertension and hyperaldosteronism.

The present invention concerns a series of novel peptides which inhibitrenin. It also concerns pharmaceutical compositions containing thesenovel peptides, methods of treating renin-associated hypertension,congestive heart failure, and hyperaldosteronism, as well as the use ofthe peptides as diagnostic tools, and the methods for preparing thepeptides.

Structurally the compounds of the instant invention represent a newclass of peptides, in that they contain an amino acid bearing anα-heteroatom directly attached to the amino acid backbone The positionsof the various amino acids may be designated by reference to theoctapeptide which is the minimal angiotensinogen sequence cleaved byrenin, namely: ##STR1##

A designation for the compounds of this invention is illustrated below.The STA is considered to occupy the P₁ -P₁ ' positions. For example##STR2##

The compounds of this invention have the α-heteroatom amino acid at theP₂ position. It is surprising to find good activity in these compounds.

SUMMARY OF THE INVENTION

The present invention relates to novel peptides of the formula

    ACYL--X--Y--W--U--V                                        (I)

and the pharmaceutically acceptable acid addition salts thereof whereinACYL, X, Y, W, U, and V are as defined herein below.

The invention also includes pharmaceutical compositions comprising aneffective amount of the above peptide of formula I in admixture with apharmaceutically acceptable carrier or excipient and a method fortreating renin-associated hypertension in a patient suffering therefromcomprising administering to said patient the above pharmaceuticalcomposition in unit dosage form.

Further the invention includes a pharmaceutical composition comprisingan effective amount of a peptide of formula I above in admixture with apharmaceutically acceptable carrier or excipient, and a method fortreating hyperaldosteronism in a patient suffering therefrom comprisingadministering to said patient the above pharmaceutical composition inunit dosage form.

The invention also includes a pharmaceutical composition comprising aneffective amount of a peptide of formula I above in admixture with apharmaceutically acceptable carrier or excipient, and a method fortreating congestive heart failure in a patient suffering therefromcomprising administering to said patient the above pharmaceuticalcomposition in unit dosage form.

The present invention also includes the use of peptides of formula Iabove as diagnostic tools for the identification of cases ofhypertension due to renin excess.

The invention further includes methods for preparing peptides of formulaI above.

DETAILED DESCRIPTION

The following table provides a dictionary of the terms used in thedescription of the invention.

                                      TABLE I                                     __________________________________________________________________________    Abbreviated                                                                   Designation                                                                   __________________________________________________________________________                       Amino Acid                                                 LEU                L-Leucine                                                  D-LEU              D-Leucine                                                  STA                4( .sub.--S)-Amino-3( .sub.--S)-hydroxy-6-methylheptano                       ic acid                                                    PHSTA              4( .sub.--S)-Amino-3( .sub.--S)-hydroxy-5-phenylpentano                       ic acid                                                    CYSTA              4( .sub.--S)-Amino-3( .sub.--S)-hydroxy-5-cyclohexanepe                       ntanoic acid                                               NORSTA             3( .sub.--S)-Amino-2( --R)-hydroxy-5-methylhexanoic                           acid                                                       ILE                L-Isoleucine                                               D-ILE              D-Isoleucine                                               N-MeLEU            N-Methylleucine                                            N-MeILE            N-Methylisoleucine                                         PHE                L-Phenylalanine                                            HOMOPHE            Homophenylalanine                                          PGY                2( .sub.--S)-Aminopentanoic acid                           VAL                L-Valine                                                   NAPHTHYLALA        Naphthylalanine                                            CYCLOHEXYLALA      Cyclohexylalanine                                          TYR                L-Tyrosine                                                 O-MeTYR            O-Methyltyrosine                                           TRP                L-Tryptophane                                              ASTA               3( --R, .sub.--S),4( .sub.--S)-Diamino-6-methylheptanoi                       c acid                                                     ACYS               3( --R, .sub.--S),4( .sub.--S)-Diamino-5-cyclohexanepen                       tanoic acid                                                CHSTA              4( .sub.--S)-Amino-3( .sub.--S)-hydroxy-4-cyclohexanebu                       tanoic acid                                                DFSTA              4( .sub.--S)-Amino-3( .sub.--S)-hydroxy-2,2-difluoro-6-                       methylheptanoic acid                                       DFKSTA             4( .sub.--S)-Amino-3-oxo-2,2-difluoro-6-methylheptanoic                        acid                                                      DFCYS              4( .sub.--S)-Amino-3( .sub.--S)-hydroxy-2,2-difluoro-5-                       cyclohexanepentanoic acid                                  DFKCYS             4( .sub.--S)-Amino-3-oxo-2,2-difluoro-5-cyclohexanepent                       anoic acid                                                 DFCHS              4( .sub.--S)-Amino-3( .sub.--S)-hydroxy-2-2-difluoro-4-                       cyclohexanebutanoic acid                                   DFKCHS             4( .sub.--S)-Amino-3-oxo-2,2-difluoro-4-cyclohexanebuta                       noic acid                                                                     Acyl Group                                                 TOS                p-Toluenesulfonyl                                          PHT                Phthaloyl                                                  Z                  Benzyloxycarbonyl                                          BOC                Tert-butyloxycarbonyl                                      DNMA               Di-(1-naphthylmethyl)acetyl                                BMA                3-Amino-3-methylbutanoyl                                   Z-BMA              3-(Benzyloxycarbonylamino)-3-methylbutanoyl                BBSP               2-Benzyl-3-(t-butylsulfonyl)propionyl                      IVA                Isovaleryl                                                 NVA                n-Valeryl                                                                     Amides With                                                NHCH.sub.2 Ph      Benzylamine                                                 ##STR3##          Cyclohexylmethylamine                                       ##STR4##          m-Xylene-di-amine (Z or BOC)                                ##STR5##          m-Xylene-di-amine                                          NH.sub.2           Ammonia                                                     ##STR6##          4-Amino-N-benzyl-piperidine                                 ##STR7##          4-Aminopiperidine                                           ##STR8##          2-Aminomethylpyridine                                      NHCH.sub.2 CH(CH.sub.3)CH.sub.2 CH.sub.3                                                         2-Methylbutylamine                                          ##STR9##          1-Hydroxymethyl-2-methyl-butylamine                         ##STR10##         4-(2-Aminoethyl)morpholine                                                    Protecting Group                                           Z                  Benzyloxycarbonyl                                          BOC                Tert-butyloxycarbonyl                                                         Esters With                                                OCH.sub.3          Methanol                                                   OC.sub.2 H.sub.5   Ethanol                                                    O-t-Bu             t-Butanol                                                  O-i-Pr             Isopropanol                                                                   Solvents and Reagents                                      DMF                N,N-Dimethylformamide                                      HOBT.H.sub.2 O     Hydroxybenzotriazole hydrate                               DCC                N,N'-Dicyclohexylcarbodiimide                              HOAc               Acetic acid                                                Et.sub.3 N         Triethylamine                                              THF                Tetrahydrofuran                                            EtOH               Ethanol                                                    MeOH               Methanol                                                   Et.sub.2 O         Diethylether                                               EtOAc              Ethyl acetate                                              __________________________________________________________________________

The peptides of the present invention are represented by the formula

    ACYL--X--Y--W--U--V                                        (I)

or a pharmaceutically acceptable acid addition salt thereof, wherein##STR11## wherein R and R' are each independently hydrogen or straightor branched chain lower alkyl, ##STR12## is a saturated ring containing1 to 5 carbon atoms wherein Q is CH₂, O, S, or NR;

X is absent, PHE, HOMOPHE, NATHTHYLALA, CYCLOHEXYLALA, O-MeTYR, TYR, orTRP, with the proviso that when ACYL is DNMA, BBSP, or ##STR13##

X is absent.

Y is ##STR14## wherein R₁ is lower alkyl, lower alkenyl, lower alkynyl,aryl, heteroaryl, aralkyl,

(CH₂)_(n) -NHR₂, wherein n is an integer of from 2 to 4, and R₂ is##STR15## wherein R₆ is hydrogen, lower alkyl or aryl, wherein R₇ is R₁,##STR16## wherein R₈ is lower alkyl or together with R₇, when R₇ islower alkyl, forms a heterocyclic ring containing from 4 to 6 carbonatoms optionally containing one or more S, O or NR; R₉ is alkyl oraralkyl;

W is STA, PHSTA, CYSTA, ASTA, ACYS, CHSTA, DFSTA, DFKSTA, DFCYS, DFKCYS,DFCHS, DFKCHS, or NORSTA;

U is absent, LEU, ILE, VAL, N-MeLEU, N-MeILE; and V is ##STR17##

Preferred compounds of the present invention are those of formula Iwherein in the Y fragment, the meaning of R₁ and R₇ are methyl, ethyl,isopropyl, propyl, allyl, propargyl, penyl, benzyl, ##STR18## or where nis an integer of from 2 to 4 and R₂ is ##STR19## wherein R₃ is H ormethyl, ##STR20## wherein R₅ is ##STR21## wherein R₆ is H or methyl.

Also preferred are compounds of formula I wherein R₇ is H, ##STR22##where R₆ is H or methyl, ##STR23## wherein R₅ is ##STR24## wherein R₃ isH or alkyl.

Also preferred are compounds according to claim 1 wherein in Y, R₇ andR₈ are each independently lower alkyl, or together form ##STR25##

Other preferred compounds of the instant invention are represented byformula I wherein ACYL is BOC, IVA, DNMA, BMA, BBSP, or ##STR26##

Other preferred compounds of the instant invention are those of formulaI wherein W is STA, CYSTA, CHSTA, ACYS, DFKCYS, or DFKST.

Still other preferred compounds of the instant invention those offormula I wherein V is ##STR27##

Particularly valuable compounds falling within the scope of the instantinvention include the following: ##STR28##

The compounds include solvates and hydrates and pharmaceuticallyacceptable acid addition salts of the basic compounds of formula Iabove.

The term pharmaceutically acceptable acid addition salt is intended tomean a relatively nontoxic acid addition salt either from inorganic ororganic acids such as, for example, hydrochloric, hydrobromic,hydroiodic, sulfuric, phosphoric, acetic, citric, oxalic, malonic,salicylic, malic, benzoic, gluconic, fumaric, succinic, ascorbic,maleic, tartaric, methanesulfonic and the like. The salts are preparedby contacting the free base form with a sufficient amount of the desiredacid to produce a salt in the conventional manner. The free base formsmay be regenerated by treating the salt form with a base.

The modified peptides of the present invention possess one or morechiral centers and each center may exist in the R(D) or S(L)configuration. The present invention includes all enantiomeric andepimeric forms as well as the appropriate mixtures thereof.

Novel intermediates of the present invention include the followingcompounds:

DNMA-NHCH(OCH₃)CO₂ CH₃,

DNMA-NHCH(OCH₃)CO₂ H,

DNMA-NHCH(SCH₃)CO₂ H,

DNMA-NHCH(SCH(CH₃)₂)CO₂ H,

DNMA-NHCH(OH)CO₂ H,

DNMA-NHCH(OC₂ H₅)CO₂ C₂ H₅,

DNMA-NHCH(OC₂ H₅)CO₂ H, ##STR29##

Some of the above novel peptides may be prepared in accordance withwell-known procedures for preparing peptides from their constituentamino acids. Other of the novel peptides of the present invention areprepared by a step-wise procedure or by a fragment coupling proceduredepending upon the particular final product desired.

The following schemes illustrate novel methods of preparing certainpeptides of the present invention. ##STR30##

The intermediates of Scheme I are novel and are prepared by anadaptation of a method described by U. Zollner and D. Ben-Ishai,Tetrahedron 31, 863(1975). Those compounds bearing an α-oxygen atom areprepared in the following manner. DNMA-NH₂ and glyoxylic acid hydrateare heated in acetone for 4 to 24 hours, giving the α-OH compound I. Iis dissolved in the appropriate alcohol, cooled to 0°, and treated withconcentrated H₂ SO₄. The mixture is allowed to warm to room temperaturefor 8 to 24 hours giving the corresponding ester of the α-alkoxycompound. The ester is hydrolyzed at room temperature for 2 to 24 hourswith strong bases as NaOH, KOH, or LiOH, giving the desired product.

The α-sulfur atom derivatives may also be prepared from the α-hydroxycompound I. In this procedure I is dissolved in HOAc, cooled to 0°, andtreated with concentrated H₂ SO₄ and the appropriate mercaptan. Afterstirring at room temperature for 1 to 4 days, the α-mercapto derivativecan be isolated.

The novel α-amino derivatives can be prepared according to Scheme IIIfrom the completed peptide. The peptide is dissolved in THF, cooled to-60° and treated with mercuric chloride. The appropriate amine is thenadded and the mixture is allowed to warm to room temperature over 4 to24 hours, giving the desired product. This procedure is an adaptation ofthat described by M. G. Bock, R. M. DiPardo, and R. Freidinger, J. Org.Chem. 51, 3718 (1986).

Scheme II describes preparing the completed peptide. The appropriateDNMA acylated α-heteroatom amino acid derivative is coupled to STA-NHCH₂CH(CH₃)CH₂ CH₃ in an inert solvent such as DMF, THF, CHCl₃, CH₂ Cl₂, orEtOAc at -5° to 25° for 4 to 24 hours, in the presence of DCC and HOBT.

α-Oxygen-containing compounds of formula I are prepared by:

(a) reacting DNMA-NH₂ with glyoxylic acid to produce DNMA-NHCH(OH)CO₂ H,

(b) reacting the product of step (a) with an alcohol to produceDNMA-NHCH(OR₁)CO₂ R₁, wherein R₁ is lower alkyl, lower alkenyl, loweralkynyl, aryl, heteroaryl, aralkyl, (CH₂)_(n) -NHR₂, wherein n is aninteger of from 2 to 4, and R₂ is ##STR31## wherein R₃ is is hydrogen,lower alkyl, or aryl, R₄ is H, lower alkyl or aralkyl, R₅ is ##STR32##wherein R₆ is hydrogen, lower alkyl or aryl, (c) hydrolyzing the productof step (b) to DNMA-NHCH(OR₁)CO₂ H wherein R₁ is as defined above, and

(d) reacting the product of step (c) with the desired fragment toproduce a peptide of formula I, and converting, if desired, to apharmaceutically acceptable salt thereof.

α-Sulphur-containing compounds of formula I are prepared by:

(a) reacting DNMA-NH₂ with glyoxylic acid to produce DNMA-NHCH(OH)CO₂ H,

(b) reacting the product of step (a) with a mercaptan to produceDNMA-NHCH(SR₁)CO₂ H wherein R₁ is lower alkyl, lower alkyenyl, loweralkynyl, aryl, heteroaryl, aralkyl, (CH₂)_(n) -NHR₂, wherein n is aninteger of from 2 to 4, and R₂ is ##STR33## wherein R₃ is hydrogen,lower alkyl, or aryl, R₄ is H, lower alkyl or aralkyl, R₅ is ##STR34##wherein R₆ is hydrogen, lower alkyl or aryl, (c) coupling the product ofstep (b) with STA-NHCH₂ CH(CH₃)CH₂ CH₃ to produce the correspondingDNMA-NHCH(SR₁)CO-STA-NHCH₂ CH(CH₃)CH₂ CH₃ wherein R₁ is as above, andconverting, if desired, to a pharmaceutically acceptable salt thereof.

α-Amino-containing compounds of formula I are prepared by treatingDNMA-NHCH(SR₁)CO-STA-NHCH₂ CH(CH₃)CH₂ CH₃ with mercuric chloride in thepresence of an amine to produce a desired compound of formula I andconverting, if desired, to a pharmaceutically acceptable salt thereof.

Using BBSP-NH₂, the amide derived from2-benzyl-3-(t-butylsulfonyl)propionic acid (EP-236,734) in place ofDNMA-NH₂ and following the routes outlined in Schemes I, II, and III,certain peptides of the present invention can be prepared. Likewise,using Z-BMA-NH₂, the amide derived from3-(benzyloxycarbonylamino)-3-methylbutanoic acid (EP-229,667), andfollowing the routes outlined in Schemes I, II, and III, certainprecursors of other target peptides of the present invention can beprepared. Removal of the benzyloxycarbonyl group gives certain otherpeptides of the present invention.

The strategy of peptide chain assembly and selection and removal ofprotecting groups is discussed in Chapter 1, "The Peptide Bond", in "ThePeptides. Analysis, Synthesis, Biology," E. Gross and J. Meienhofer,Eds., Academic Press, New York, N.Y., 1979, Vol. 1, pp. 42-44.

The DCC/HOBT method of coupling is well known to those skilled in theart and is discussed in Chapter 5, "The Carbodiimide Method" by D. H.Rich and J. Singh in "The Peptides. Analysis, Synthesis, Biology," E.Gross and J. Meienhofer, Eds., Academic Press, New York, NY, 1979, Vol.1, pp. 241-261.

Peptide coupling depends on activating the carboxyl group of theprotected amino acid prior to condensing it with another peptidecontaining a free amino terminus. In addition to the DCC coupling methoddescribed above, other methods of activating the carboxyl group of aprotected amino acid include:

1) The azide method - described in Chapter 4 of the above reference.

2) The mixed anhydride method - described in Chapter 6 of the abovereference.

3) The active ester method - described in Chapter 3 of the abovereference.

The acyl groups derived from the substituted succinic acid amides may beprepared as follows. 1-Naphthaldehyde is reacted with diethyl succinatein a Stobbe condensation, and the corresponding di-acid is converted tothe anhydride with acetic anhydride. Treatment with the appropriateamine gives 2-(1-naphthylmethylene)-3-(substituted aminocarbonyl)propionic acid. Catalytic hydrogenation gives the desired2-(1-naphthylmethyl)-3-(substituted aminocarbonyl)propionic acid. Thisacid may be condensed with suitably protected amino acids using thecoupling methods known to peptide chemistry, for example, thecarbodiimide method. This is discussed in European ApplicationPublication No. 206,807 and European Application Publication No.200,406.

The compounds of the present invention are useful for treatingrenin-associated hypertension, congestive heart failure, andhyperaldosteronism. They are also useful as diagnostic tools fordetermining the presence of renin-associated hypertension orhyperaldosteronism.

The term lower alkyl refers to straight or branched chain hydrocarbonradicals containing from 1 to 10 carbon atoms including but not limitedto methyl, ethyl, n-propyl, isopropyl, n-butyl, iso-butyl, sec-butyl,2-methylhexyl, n-pentyl, 1-methylbutyl, 2,2-dimethylbutyl,2-methylpentyl, 2,2-dimethylpropyl, n-hexyl and the like.

Lower alkenyl means a straight or branched chain hydrocarbon radical offrom 1 to 10 carbon atoms containing a double bond. This includes but isnot limited to allyl and methylallyl.

Lower alkynyl means a straight or branched chain hydrocarbon radical offrom 1 to 10 carbon atoms containing a triple bond. This includes but isnot limited to propargyl.

Aryl means phenyl, naphthyl or other aromatic groups, including mono- orbicyclic, which may be substituted, especially monosubstituted, by F,Cl, Br, I, CF₃, OH, OR, or R, wherein R is lower alkyl.

Heteroaryl means aromatic heterocyclic rings containing at least oneheteroatom selected from O, S, and N and from 3 to 5 carbon atomsincluding but not limited to thiazoles and imidazoles.

Aralkyl is as described above for alkyl and aryl, including but notlimited to benzyl.

Substituted alkyl includes but is not limited to such groups as hydroxyand halogen.

Pharmaceutical compositions which comprise an effective amount of thecompound in combination with a pharmaceutically acceptable carrier arepart of the present invention. An important aspect of the presentinvention is a method of treating renin-associated hypertension in amammal which comprises administering a pharmaceutical compositioncontaining an effective amount of a compound of the invention incombination with a pharmaceutically acceptable carrier to the mammal.

Another equally important aspect of the present invention is a method oftreating hyperaldosteronism in a mammal which comprises administering apharmaceutical composition containing an effective amount of a compoundof the invention in combination with a pharmaceutically acceptablecarrier to the mammal.

Yet another important aspect of the present invention is method oftreating congestive heart failure in a mammal which comprisesadministering a pharmaceutical composition containing an effectiveamount of a compound of the invention in combination with apharmaceutically acceptable carrier.

The effectiveness of the aforementioned compounds is determined by atest for in vitro renin inhibitory activity. This activity is determinedby a standard radioimmunoassay for angiotensin I. In this assay theenzyme, renin, incubated for 2 hours at 37° in the presense of asubstrate, angiotensinogen, generates the product, angiotensin I. Testcompounds are added to the incubation mixture. Relative activity isreported as the percent inhibition at 10⁻⁶ molar concentration.

                                      TABLE II                                    __________________________________________________________________________                                             Activity                             Inhibition                               %                                    Compound                                 @ 10.sup.-6 M                        __________________________________________________________________________    DNMANHCH(OCH.sub.3)COSTANHCH.sub.2 CH(CH.sub.3)CH.sub.2 CH.sub.3                                                       90                                   DNMANHCH(SCH.sub.3)COSTANHCH.sub.2 CH(CH.sub.3)CH.sub.2 CH.sub.3                                                       86                                   DNMANHCH(SOCH.sub.3)COSTANHCH.sub.2 CH(CH.sub.3)CH.sub.2 CH.sub.3                                                      35                                   DNMANHCH(SCH(CH.sub.3).sub.2)COSTANHCH.sub.2 CH(CH.sub.3)CH.sub.2             CH.sub.3                                 87                                   (fast isomer)                                                                 DNMANHCH(SCH(CH.sub.3).sub.2)COSTANHCH.sub.2 CH(CH.sub.3)CH.sub.2             CH.sub.3                                 26                                   (slow isomer)                                                                 DNMANHCH(NH.sub.2)COSTANHCH.sub.2 CH(CH.sub.3)CH.sub.2 CH.sub.3                                                         9                                   (fast isomer)                                                                 DNMANHCH(NH.sub.2)COSTANHCH.sub.2 CH(CH.sub.3)CH.sub.2 CH.sub.3                                                        54                                   (slow isomer)                                                                 DNMANHCH(NHCO.sub.2 Et)COSTANHCH.sub.2 CH(CH.sub.3)CH.sub.2 CH.sub.3                                                   40                                   DNMANHCH(OC.sub.2 H.sub.5)COSTANHCH.sub.2 CH(CH.sub.3)CH.sub.2 CH.sub.3                                                100                                  (fast isomer)                                                                 DNMANHCH(OC.sub.2 H.sub.5)COSTANHCH.sub.2 CH(CH.sub.3)CH.sub.2 CH.sub.3                                                15                                   (slow isomer)                                                                 DNMANHCH(NH(CH.sub.2).sub.3 NHCSNHCH.sub.3)COSTANHCH.sub.2 CH(CH.sub.3)CH.    sub.2 CH.sub.3                           36                                   DNMANHCH(NHCOCH.sub.3)COSTANHCH.sub.2 CH(CH.sub.3)CH.sub.2 CH.sub.3                                                    39                                   DNMANHCH(NHPh)COSTANHCH.sub.2 CH(CH.sub.3)CH.sub.2 CH.sub.3                                                            56                                   DNMANHCH(SPh)COSTANHCH.sub.2 CH(CH.sub.3)CH.sub.2 CH.sub.3                                                             40                                   DNMANHCH(N(CH.sub.3).sub.2)COSTANHCH.sub.2 CH(CH.sub.3)CH.sub.2 CH.sub.3                                               55                                   (slow isomer)                                                                 DNMA-NHCH(NHCH(CH.sub.3).sub.2)COSTANHCH.sub.2 CH(CH.sub.3)CH.sub.2           CH.sub.3                                 60                                   (slow isomer)                                                                 DNMANHCH(NHC.sub.2 H.sub.5)COSTANHCH.sub.2 CH(CH.sub.3)CH.sub.2 CH.sub.3                                               71                                   (slow isomer)                                                                  ##STR35##                               57                                    ##STR36##                               100                                  __________________________________________________________________________

As can be seen from the above table, the compounds of the presentinvention have a significant effect on the activity of renin and thusare useful for the treatment of hypertension, congestive heart failure,and hyperaldosteronism.

For preparing pharmaceutical compositions from the compounds describedby this invention, inert, pharmaceutically acceptable carriers can beeither solid or liquid. Solid form preparations include powders,tablets, dispersible granules, capsules, cachets, and suppositories. Asolid carrier can be one or more substances which may also act asdiluents, flavoring agents, solubilizers, lubricants, suspending agents,binders or tablet disintegrating agents; it can also be encapsulatingmaterial. In powders, the carrier is a finely divided solid which is inadmixture with the finely divided active compound. In the tablet theactive compound is mixed with carrier having the necessary bindingproperties in suitable proportions and compacted in the shape and sizedesired. The powders and tablets preferably contain from 5 to 10 toabout 70 percent of the active ingredient. Suitable solid carriers aremagnesium carbonate, magnesium stearate, talc, sugar, lactose, pectin,dextrin, starch, gelatin, tragacanth, methylcellulose, a low meltingwax, cocoa butter, and the like. The term "preparation" is intended toinclude the formulation of the active compound with encapsulatingmaterial as carrier providing a capsule in which the active component(with or without other carriers) is surrounded by carrier, which is thusin association with it. Similarly, cachets are included. Tablets,powders, cachets, and capsules can be used as solid dosage formssuitable for oral administration.

The compounds of the present invention may be administered orally,buccally, parenterally, by inhalation spray, rectally or topically indosage unit formulations containing conventional nontoxicpharmaceutically acceptable carriers, adjuvants and vehicles as desired.The term parenteral as used herein includes subcutaneous injections,intravenous, intramuscular, intrasternal injection or infusiontechniques.

For preparing suppositories, a low melting wax such as a mixture offatty acid glycerides or cocoa butter is first melted, and the activeingredient is dispersed homogeneously therein by stirring. The moltenhomogeneous mixture is then poured into conventient sized molds, allowedto cool, and thereby solidify.

Liquid form preparations include solutions, suspensions, and emulsions.As an example may be mentioned water or water/propylene glycol solutionsfor parenteral injection. Liquid preparations can also be formulated insolution in aqueous polyethyleneglycol solution. Aqueous suspensionssuitable for oral use can be made by dispersing the finely dividedactive component in water with viscous material, i.e., natural orsynthetic gums, resins, methylcellulose, sodium carboxymethylcellulose,and other well-known suspending agents.

Also included are solid form preparations which are intended to beconverted, shortly before use, to liquid form preparations for eitheroral or parenteral administration. Such liquid forms include solutions,suspensions, and emulsions. These particular solid form preparations aremost conveniently provided in unit dose form and as such are used toprovide a single liquid dosage unit. Alternately, sufficient solid maybe provided so that after conversion to liquid form, multiple individualliquid doses may be obtained by measuring predetermined volumes of theliquid form preparation as with a syringe, teaspoon, or other volumetriccontainer. When multiple liquid doses are so prepared, it is preferredto maintain the unused portion of said liquid doses at low temperature(i.e., under refrigeration) in order to retard possible decomposition.The solid form preparations intended to be converted to liquid form maycontain, in addition to the active material, flavorants, colorants,stabilizers, buffers, artificial and natural sweeteners, dispersants,thickeners, solubilizing agents, and the like. The liquid utilized forpreparing the liquid form preparation may be water, isotonic water,ethanol, glycerin, propylene glycol, and the like, as well as mixturesthereof. Naturally, the liquid utilized will be chosen with regard tothe route of administration, for example, liquid preparations containinglarge amounts of ethanol are not suitable for parenteral use.

Preferably, the pharmaceutical preparation is in unit dosage form. Insuch form, the preparation is subdivided into unit doses containingappropriate quantities of the active component. The unit dosage form canbe a packaged preparation, the package containing discrete quantities ofpreparation, for example, packeted tablets, capsules, and powders invials or ampules. The unit dosage form can also be a capsule, cachet, ortablet itself, or it can be the appropriate number of any of these inpackaged form.

The quantity of active compound in a unit dose of preparation may bevaried or adjusted from 1 mg to 500 mg, preferably 5 to 100 mg accordingto the particular application and the potency of the active ingredient.The compositions can, if desired, also contain other compatibletherapeutic agents.

In therapeutic use as renin inhibitors, the mammalian dosage range for a70 kg subject is from 1 to 1500 mg per day or preferably 25 to 750 mgper day optionally in divided portions. The dosages, however, may bevaried depending upon the requirements of the patient, the severity ofthe condition being treated and the compound being employed.Determination of the proper dosage for a particular situation is withinthe skill of the art. Generally, treatment is initiated with smallerdosages which are less than the optimum dose of the compound. Thereafterthe dosage is increased by small increments until the optimum effectunder the circumstances is reached. For convenience, the total dailydosage may be divided and administered in portions during the day ifdesired.

The following examples are provided to enable one skilled in the art topractice the present invention. These examples are not intended in anyway to limit the scope of the invention but are illustrative thereof.

EXAMPLE 1 DNMA-NHCH(OCH₃)CO-STA-NHCH₂ CH(CH₃)CH₂ CH₃

DNMA-NHCH(OCH₃)CO₂ H (1.30 g, 3.04 mmole) was dissolved in 30 ml CH₂ Cl₂and cooled to 0°. A solution of HOBT.H₂ O (0.42 g, 3.13 mmole) in 3 mlDMF was added, followed by DCC (0.65 g, 3.13 mmole). A solution ofSTA-NHCH₂ CH(CH₃)CH₂ CH₃ (0.74 g, 3.04 mmole) in 15 ml CH₂ Cl₂ wasadded, followed by stirring overnight at 23°. The resulting suspensionwas filtered and the filtrate evaporated to an oil and taken up inEtOAc. The solution was washed with 1N citric acid, saturated NaCl,saturated NaHCO₃, and saturated NaCl, followed by drying over MgSO₄. Thesolution was evaporated under reduced pressure to a foam which waschromatographed on silica gel, eluting with EtOAc/CHCl₃ (1/1). Theproduct was obtained as a white foam, 1.5 g, 75.4% yield. The structurewas confirmed by NMR and mass spectroscopy.

Calcd. for C₄₀ H₅₁ N₃ O₅ (MW 653.87):

C, 73.48; H, 7.86; N, 6.43;

Found: C, 73.34; H, 7.94; N, 6.35.

EXAMPLE 2 DNMA-NHCH(SCH₃)CO-STA-NHCH₂ CH(CH₃)CH₂ CH₃

DNMA-NHCH(SCH₃)CO₂ H (5.86 g, 13.2 mmole) was dissolved in 100 ml CH₂Cl₂ and cooled to 0°. A solution of HOBT.H₂ O (1.84 g, 13.6 mmole) in 6ml DMF was added. DCC (2.81 g, 13.6 mmole) was added, followed by asolution of STA-NHCH₂ CH(CH₃)CH₂ CH₃ (3.22 g, 13.2 mmole) in 25 ml ofCH₂ Cl₂. After stirring at 23° overnight, the mixture was filtered, andthe filtrate evaporated to an oil. The oil was dissolved in EtOAc, thesolution filtered, and the filtrate washed with 1N citric acid,saturated NaCl, saturated NaHCO₃, and saturated NaCl. The organic phasewas dried over MgSO₄, and evaporated under reduced pressure to a whitefoam. The foam was chromatographed on silica gel, eluting withEtOAc/CHCl₃ (1/1), giving the product as a white foam, 7.37 g, 83%yield. The structure was confirmed by NMR and mass spectroscopy.

Calcd. for C₄₀ H₅₁ N₃ O₄ S.0.125 H₂ O (MW 672.19):

C, 71.47; H, 7.68; N, 6.25; H₂₀, 0.33; S, 4.77;

Found: C, 71.16; H, 7.55; N, 6.21; H 0, 0.33; S, 4.72.

EXAMPLE 3 DNMA-NHCH(SOCH₃)CO-STA-NHCH₂ CH(CH₃)CH₂ CH₃

DNMA-NHCH(SCH₃)CO-STA-NHCH₂ CH(CH₃)CH₂ CH₃ (1.5 g, 2.24 mmole) wasdissolved in 25 ml MeOH, to which was added a solution of NaIO₄ (1.01 g,4.68 mmole) in 10 ml H₂ O. After warming to 50° for 2 hours, the mixturewas evaporated to an oil. The oil was taken into EtOAc and extractedwith H₂ O, 10% sodium bisulfite solution, saturated NaHCO₃ and saturatedNaCl. After drying over MgSO₄, the mixture was evaporated under reducedpressure to a foam. The foam was chromatographed on silica gel, elutingwith EtOAc/CHCl₃ (3/1). Combination of the appropriate factions gave theproduct as a white foam, 0.85 g. The structure was confirmed by NMR andmass spectroscopy.

Calcd. for C₄₀ H₅₁ N₃ O₅ S (MW 685.93):

C, 70.04; H, 7.49; N, 6.13; S, 4.67;

Found: C, 70.06; H, 7.54; N, 6.03; S, 4.51

EXAMPLES 4 AND 5 DNMA-NHCH(SCH(CH₃)₂)CO-STA-NHCH₂ CH(CH₃)CH₂ CH₃

DNMA-NHCH(SCH(CH₃)₂)CO₂ H (7.48 g, 15.9 mmole) was dissolved in 130 mlCH₂ Cl₂. A solution of HOBT.H₂ O (2.2 g, 16.3 mmole) in 4 ml DMF wasadded, and the mixture was cooled to 0°. DCC (3.37 g, 16.3 mmole) wasadded, followed by a solution of STA-NHCH₂ CH(CH₃)CH₂ CH₃ (3.87 g, 15.9mmole) in 25 ml CH₂ Cl₂. The mixture was stirred at 23° overnight,filtered, and the filtrate evaporated to an oil, which was taken up inEtOAc and filtered. The filtrate was washed with 1N citric acid,saturated NaCl, saturated NaHCO₃, and saturated NaCl. The solution wasdried over MgSO₄ and evaporated under reduced pressure to a foam whichwas chromatographed on silica gel, eluting with EtOAc/CHCl₃ (1/1).Fractions containing the faster eluting isomer were combined, giving theproduct as a white foam, 1.91 g. The structure was confirmed by NMR andmass spectroscopy.

Calcd. for C₄₂ H₅₅ N₃ O₄ S. 0.06 CHCl₃ (MW 705.45):

C, 71.61; H, 7.86; N, 5.95; S, 4.54; Cl, 0.94

Found: C, 71.53; H, 8.13; N, 6.18; S, 4.40; Cl, 0.74

Fractions containing the slower eluting diastereomer were combinedgiving 1.80 g of the product as a foam, identified by NMR and massspectroscopy.

Calcd. for C₄₂ H₅₅ N₃ O₄ S. 0.5 H₂ O (MW 706.97):

C, 71.35; H, 7.98; N, 5.94; S, 4.53; H₂ O, 1.27;

Found: C, 71.50; H, 7.83; N, 5.91; S, 4.68; H₂ O, 1.01 .

EXAMPLES 6 AND 7 DNMA-NHCH(NH₂)CO-STA-NHCH₂ CH(CH₃)CH₂ CH₃

DNMA-NHCH(SCH₃)CO-STA-NHCH₂ CH(CH₃)CH₂ CH₃ (2.91 g, 4.34 mmole) wasdissolved in 100 ml THF, and mercuric chloride (1.76 g, 6.48 mmole) wasadded. After cooling to -60°, the mixture was saturated with anhydrousNH₃ gas. After warming to 23° overnight, the mixture was recooled andsaturated with NH₃ gas again, and allowed to warm to 23° over 4 hours.The mixture was evaporated to a foam, taken into Et₂ O and filtered. Thefiltrate was briefly purged with HCl gas until acidic to wet litmus. Awhite solid precipitated, which was filtered, washed with Et₂ O, dried,and chromatographed on silica gel, eluting with a gradient of 0-5% MeOHin CHCl₃. Combination of the fractions containing the faster elutingisomer yielded a foam which was dissolved in Et₂ O and purged brieflywith HCl gas. The resulting solid was filtered, washed with Et₂ O anddried giving the product as a white solid, 0.87 g. The structure wasconfirmed by NMR and mass spectroscopy.

Calcd. for C₃₉ H₅₀ N₄ O₄. 1.1 HCl.0.5 H₂ O (MW 687.97):

C, 68.09; H, 7.63; N, 8.14; Cl, 5.67; H₂ O, 1.31 ;

Found: C, 68.50; H, 7.65; N, 8.06; Cl, 5.92; H₂ O, 1.69.

Fractions containing the slower eluting diastereomer were similarlytreated, giving a white solid, 0.92 g. The structure was confirmed byNMR and mass spectroscopy.

Calcd. for C₃₉ H₅₀ N₄ O₄.1.2 HCl.0.6 H₂ O (MN 693.42):

C, 67.55; H, 7.61; N, 8.08; Cl, 6.13; H₂ O, 1.56;

Found: C, 68.37; H, 7.78; N, 8.07; Cl, 6.33; H₂ O, 1.53.

Fractions containing a mixture both isomers were evaporated to a whitefoam, 0.35 g, which was used in the preparation of Example 8.

EXAMPLE 8 DNMA-NHCH(NHCO₂ Et)CO-STA-NHCH₂ CH(CH₃)CH₂ CH₃

DNMA-NHCH(NH₂)CO-STA-NHCH₂ CH(CH₃)CH₂ CH₃ (0.35 g, 0.5 mmole) as amixture of diastereomers, was dissolved in 20 ml THF. Et₃ N (0.078 ml,0.56 mmole) was added, followed by ethyl chloroformate (0.055 ml, 0.56mmole). After stirring 2 hours at 23°, the mixture was filtered, thecollected solid washed with THF, and the filtrate evaporated to a foam.The residue was taken up in EtOAc, washed with 1N citric acid, saturatedNaCl, saturated NaHCO₃, and saturated NaCl. The organic phase was driedover MgSO₄ and evaporated under reduced pressure giving the product as afoam, 0.38 g. The structure was confirmed by NMR and mass spectroscopy.

Calcd. for C₄₂ H₅₄ N₄ O₆ (MW 710.92):

C, 70.96; H, 7.65; N, 7.88;

Found C, 71.22; H, 7.78; N, 7.92.

EXAMPLES 9 AND 10 DNMA-NHCH(OC₂ H₅)CO-STA-NHCH₂ CH(CH₃)CH₂ CH₃

DNMA-NHCH(OC₂ H₅)CO₂ H (1.20 g, 2.72 mmole) was dissolved in 50 ml CH₂Cl₂ A solution of HOBT.H₂ O(0.38 g, 2.8 mmole) in 4 ml DMF was added,followed by cooling the mixture to 0°. STA-NHCH₂ CH(CH₃)CH₂ CH₃ (0.67 g,2.72 mmole) dissolved in 20 ml CH₂ Cl₂ was added, followed by theaddition of DCC (0.58 g, 2.8 mmole). The mixture was stirred and allowedto warm to 23° overnight. The mixture was filtered, evaporated to asyrup, and taken up into EtOAc. The mixture was again filtered, andwashed with 1N citric acid, saturated NaCl, saturated NaHCO₃, andsaturated NaCl. The mixture was dried over MgSO₄, filtered, andevaporated to a foam, 1.86 g. The foam was chromatographed on silica geleluting with EtOAc/CHCl₃ (1/1). Combination of fractions containing thefaster eluting isomer gave a foam which was recrystallized from Et₂ Ogiving 0.45 g of the product as a solid. The structure was confirmed byNMR and mass spectroscopy.

Calcd. for C₄₁ H₅₃ N₃ O₅ (MW 667.90):

C, 73.73; H, 7.99; N, 6.29;

Found: C, 73.57; H, 7.89; N, 6.38.

The slower eluting isomer was similarly obtained as a crystalline solid,0.48 g. The structure was confirmed by NMR and mass spectroscopy.

Calcd. for C₄₁ H₅₃ N₃ O₅ (MW 667.90):

C, 73.73; H, 7.99; N, 6.29;

Found: C, 73.73; H, 7.98; N, 6.35.

EXAMPLE 11 DNMA-NHCH(NH(CH₂)₃ NHCSNHCH₃)CO-STA-NHCH₂ CH(CH₃)CH₂ CH₃

DNMA-NHCH(NH(CH₂)₃ NH₂)CO-STA-NHCH₂ CH(CH₃)CH₂ CH₃ (2.97 g, 4.35 mmole)was dissolved in 75 ml CH₂ Cl₂ and methylisothiocyanate (0.32 g, 4.44mmole) was added. After stirring at 23° overnight, the mixture wasevaporated to a foam and dissolved in EtOAc. The solution was washedwith 1N HCl, saturated NaCl, 1N NaOH, and saturated NaCl. The solutionwas dried over MgSO₄ and evaporated under reduced pressure to give thecrude product as a white foam, 3.13 g. The crude product waschromatographed on silica gel eluting with CHCl₃ /EtOAc (40/60) with agradient to 15% MeOH. The product was recovered as a white foam, 2.41 g.Trituration with Et₂ O gave 1.9 g of a white solid. The structure wasconfirmed by NMR and mass spectroscopy.

Calcd. for C₄₄ H₆₀ N₆ O₄ S. 0.75H₂ O. 0.08CHCl₃ (MW 792.13):

C, 66.83; H, 7.83; N, 10.61; S, 4.05; H₂ O, 1.70; Cl, 1.07;

Found: C, 67.77; H, 7.83; N, 10.40; S, 4.13; H₂ O, 1.69; Cl, 1.24.

EXAMPLE 12 DNMA-NHCH(NHCOCH₃)CO-STA-NHCH₂ CH(CH₃)CH₂ CH₃

DNMA-NHCH(NH₂)CO-STA-NHCH₂ CH(CH₃)CH₂ CH₃.HCl (0.73 g, 1.05 mmole) ofthe slow isomer (Example 7) was suspended in 25 ml of CH₂ Cl₂ andtreated with 0.35 ml (1.08 mmole) of Et₃ N causing solution. This wasthen treated with 0.09 ml (1.08 mmole) of acetyl chloride and stirred atroom temperature for 2.5 hours. The mixture was filtered and the solventremoved under reduced pressure. The solid residue was taken up in EtOAcand washed with H₂ O, 1N citric acid, and saturated NaCl. Drying andremoval of the solvent under reduced pressure gave a paste which wastriturated with Et₂ O giving 0.62 g of the product as a white powder.The structure was confirmed by NMR and mass spectroscopy.

Calcd. for C₄₁ H₅₂ N₄ O₅ (MW 680.89):

C, 72.32; H, 7.70; N, 8.23;

Found: C, 72.08; H, 7.63; N, 825.

EXAMPLE 13 DNMA-NHCH(NHPh)CO-STA-NHCH₂ CH(CH₃)CH₂ CH₃

DNMA-NHCH(SCH(CH₃)₂)CO-STA-NHCH₂ CH(CH₃)CH₂ CH₃, a mixture ofdiastereomers, (2.20 g, 3.15 mmole) was dissolved in 100 ml dry THF andaniline (0.88 g, 9.45 mmole) was added. HgCl₂ (1.3 g, 4.75 mmole) wasadded and the solution was stirred at 23° overnight. An additionalamount of HgCl₂ was added (0.48 g, 5.15 mmole) followed by stirring at23° for two days. The mixture was filtered and the solvent evaporatedleaving a solid. This was mixed with EtOAc, filtered, and the filtratewashed with 1N HCl, saturated NaCl, 1N NaOH, and saturated NaCl. TheEtOAc phase was dried over MgSO₄, and evaporated under reduced pressureto a yellow foam, 2.23 g. Chromatography on silica gel, eluting withEtOAc/CHCl₃ (1/1) gave 1.85 g of the product as a white foam. Thestructure was confirmed by NMR and mass spectroscopy.

Calcd for C₄₅ H₅₄ N₄ O₄.0.25EtOAc.0.25H₂ O (MW 741.49):

C, 74.51; H, 7.68; N, 7.56; H₂ O, 0.61;

Found: C, 74.25; H, 7.59; N, 7.52; H₂ O, 0.62.

EXAMPLE 14 DNMA-NHCH(SPh)CO-STA-NHCH₂ CH(CH₃) CH₂ CH₃

DNMA-NHCH(SPh)CO₂ H (1.77 g, 3.50 mmole) was dissolved in 75 ml CH₂ Cl₂and cooled to 0°. A solution of HOBT.H₂ O (0.52 g, 3.85 mmole) in 6 mlDMF was added. DCC (0.79 g, 3.85 mmole) was added, followed by asolution of STA-NHCH₂ CH(CH₃)CH₂ CH₃ (0.91 g, 3.74 mmole) in 20 ml CH₂Cl₂. After stirring at 23° overnight, the mixture was filtered, and thefiltrate evaporated to an oil. The oil was dissolved in EtOAc, thesolution filtered, and the filtrate washed with 1N citric acid,saturated NaCl, saturated NaHCO₃ and saturated NaCl. The organic phasewas dried over MgSO₄, and evaporated under reduced pressure to a whitefoam, 2.66 g. The foam was chromatographed on silica gel, eluting withEtOAc/CHCl₃ (1/1) and giving the product as a white foam, 2.0 g. Thestructure was confirmed by NMR and mass spectroscopy.

Calcd. for C₄₅ H₅₃ N₃ O₄ S. 0.25H₂ O(MW 736.51):

C, 73.39; H, 7.32; N, 5.70; S, 4.35; H₂ O, 0.61 ;

Found: C, 73.10; H, 7.48; N, 5.94; S, 4.68; H₂ O, 0.33.

EXAMPLE 15

DNMA-NHCH(N(CH₃)₂ CO-STA-NHCH₂ CH(CH₃)CH₂ CH₃

DNMA-NHCH(SCH₃)CO-STA-NHCH₂ CH(CH₃)CH₂ CH₃ (1.54 g, 2.3 mmole) wasdissolved in 50 ml dry THF, and dimethylamine (5 ml) was added. HgCl₂(0.94 g, 3.45 mmole) was added and the solution was refrigerated at 4°for two days. The excess dimethylamine was removed in vacuo, the mixturefiltered, and the solvent evaporated giving a gum. The gum wasresuspended in EtOAc, the suspension filtered, and the filtrate washedwith 1N HCl, saturated NaCl solution, and 1N NaOH. An orange precipitateformed, which was filtered off. The organic phase of the filtrate waswashed with saturated NaCl, dried over MgSO₄, and evaporated underreduced pressure to a white foam, 1.49 g. Chromatography on silica gel,eluting with CHCl₃ /EtOAc/MeOH (35/55/10) separated the mixture into afaster eluting diastereomer and a slower eluting diastereomer. Theproduct, the slower eluting isomer, was obtained as a white foam, 0.82g. The structure was confirmed by NMR and mass spectroscopy.

Calcd. for C₄₁ H₅₄ N₄ O₄ (MW 666.91):

C, 73.84; H, 8.16; N, 8.40;

Found: C, 73.59; H, 8.30; N, 8.20.

EXAMPLE 16 DNMA-NHCH(NHCH(CH₃)₂)CO-STA-NHCH₂ CH(CH₃)CH₂ CH₃

DNMA-NHCH(SCH₃)CO-STA-NHCH₂ CH(CH₃)CH₂ CH₃ (1.54 g, 2.3 mmole) wasdissolved in 50 ml dry THF, and isopropylamine (1.0 ml) was added. HgCl₂(0.94 g, 3.45 mmole) was added and solution occurred. After stirring at23° for 2 hours, a precipitate formed and the mixture was refrigeratedat 4° for two days. The mixture was filtered, evaporated to a foam andresuspended in EtOAc. The mixture was filtered, and the filtrate waswashed with 1N HCl, saturated NaCl, and 1N NaOH. An orange precipitateformed which was filtered off. The organic phase was washed withsaturated NaCl, dried over MgSO₄, and evaporated under reduced pressureto a white foam, 1.62 g. Chromatography on silica gel, eluting withCHCl₃ /EtOAc/MeOH (35/55/10), separated the mixture into faster andslower eluting diastereomers. The product, the slower elutingdiastereomer, was obtained as a crystalline solid, 0.73 g. The structurewas confirmed by NMR and mass spectroscopy.

Calcd. for C₄₁ H₅₄ N₄ O₄.0.15H₂ O (MW 669.61):

C, 73.54; H, 8.17; N, 8.37; H₂ O, 0.40;

Found: C, 73.18; H, 8.15; N, 8.33; H₂ O, 0.27.

EXAMPLE 17 DNMA-NHCH(NHC₂ H₅)CO-STA-NHCH₂ CH(CH₃) CH₂ CH₃

DNMA-NHCH(SCH₃)CO-STA-NHCH₂ CH(CH₃)CH₂ CH₃) 1.54 g, 2.3 mmole) wasdissolved in 50 ml dry THF, and ethylamine (5 ml) was added. HgCl₂ (0.94g, 3.45 mmole) was added, and solution occurred followed by solidprecipitation. After stirring at 23° overnight, the mixture wasfiltered, excess amine removed under vacuum, and the mixture filteredagain. The filtrate was evaporated under reduced pressure and theresidue was resuspended in EtOAc. The mixture was filtered and thefiltrate was washed with 1N HCl, saturated NaCl, 1N NaOH, and saturatedNaCl. The solution was dried over MgSO₄ and evaporated under reducedpressure to a white foam, 1.61 g. Chromatography on silica gel, elutingwith CHCl₃ /EtOAc/MeOH (35/55/10) separated the mixture into faster andslower eluting diastereomers. The product, the slower elutingdiastereomer, was obtained as a white foam, 0.96 g. The structure wasconfirmed by NMR and mass spectroscopy.

Calcd. for C₄₂ H₅₆ N₄ O₄.0.1CHCl₃.0.25H₂ O (MW 697.43):

C, 72.50; H, 8.19; N, 8.03; H₂ O, 0.65; Cl, 1.52;

Found: C, 72.86; H, 8.35; N, 7.85; H₂ O, 0.62; Cl, 1.54.

EXAMPLES 18 AND 19 ##STR37##

(0.41 g, 3.03 mmole) were dissolved in 50 ml CH₂ Cl₂ and cooled to 0°.DCC (0.64 g, 3.1 mmole) was added, and the mixture was allowed to warmto 23° overnight. The mixture was concentrated to 25 ml in volume underreduced pressure, and filtered to remove insoluble matter. The filtratewas evaporated, and the residue was resuspended in EtOAc. A solidprecipitated, which was again filtered. The solid thus recovered wasfound to be pure, slower eluting diastereomer, 0.41 g. The structure wasconfirmed by NMR and mass spectroscopy.

Calcd. for C₃₄ H₅₆ N₆ O₉ S (MW 724.92):

C, 56.33; H, 7.79; N, 11.59; S, 4.42;

Found: C, 56.48; H, 7.94; N, 11.66; S, 4.49.

The filtrate from removal of some of the slower moving diastereomer waswashed with saturated NaHCO₃ and saturated NaCl. Drying over MgSO₄ andremoving the solvent under reduced pressure gave a foam, 1.67 g. Thefoam was chromatographed on silica gel, eluting with a gradient of 2 to5% MeOH in CHCl₃, and giving a mixture of the faster and slower elutingdiastereomers as a foam, 1.34 g. The structure was confirmed by NMR andmass spectroscopy.

Calcd. for C₃₄ H₅₆ N₆ O₉ S.0.2CHCl₃.0.6H₂ O (MW 759.61)

C, 54.08; H, 7.61; N, 11.06; S, 4.22; Cl, 2.80; H₂ O, 1.42 ;

Found: C, 53.94; H, 7.59; N, 10.86; S, 3.93; Cl, 2.70; H₂ O, 1.42.

INTERMEDIATES FOR EXAMPLE 1 DNMA-NHCH(OCH₃)CO₂ CH₃

DNMA-NHCH(OH)CO₂ H (6.0 g, 0.0145 mole) was dissolved in 60 ml MeOH andcooled to 0°. 1.2 ml concentrated H₂ SO₄ was added, and the mixture wasallowed to warm to 23° overnight. The precipitated solid was filtered,washed with MeOH, Et₂ O, and then dried giving the product as a whitesolid, 3.30 g, 51% yield. The structure was confirmed by NMR and massspectroscopy. The material was of suitable purity for use in thefollowing step.

DNMA-NHCH(OCH₃)CO₂ H

DNMA-NHCH(OCH₃)COOCH₃ (2.41 g, 5.46 mmole) was dissolved in a mixture of15 ml MeOH and 10 ml 1,4-dioxane. 5.5 ml 1N NaOH was added, and themixture was stirred for 1 hour and evaporated to an oil under reducedpressure. The oil was suspended in Et₂ O and 5.5 ml 1N HCl was added.The phases were separated and the organic phase was washed withsaturated NaCl, dried over MgSO₄ and evaporated to a white foam, 2.6 g.The strucure was confirmed by NMR and mass spectroscopy. The materialwas of suitable purity for use in the following step.

INTERMEDIATE FOR EXAMPLE 2 DNMA-NHCH(SCH₃)CO₂ H

DNMA-NHCH(OH)CO₂ H (6.4 g, 15.4 mmole) was dissolved in 90 ml glacialacetic acid and cooled to 0°. Approximately 6 ml methyl mercaptan wasadded, followed by 10 ml concentrated H₂ SO₄. After stirring for 2 daysat 23°, the mixture was poured onto ice and extracted into EtOAc. Theorganic phase was washed with saturated NaCl and then saturated NaHCO₃until the pH of the wash was basic. The organic phase, which stillcontained the product, was washed with saturated NaCl, 1N HCl, andsaturated NaCl. The organic phase was dried over MgSO₄, and evaporatedunder reduced pressure to give a white foam, 5.94 g, 89% yield. Thestructure was confirmed by NMR and mass spectroscopy. The material wasof suitable purity for use in the following step.

INTERMEDIATE FOR EXAMPLES 5 AND 6 DNMA-NHCH(SCH(CH₃ ₂)CO₂ H

DNMA-NHCH(OH)CO₂ H (6.4 g, 15.4 mmole) was dissolved in 90 ml glacialacetic acid and cooled to 0°. To the solution was added isopropylmercaptan (6 ml, 64.6 mmole) and 10 ml concentrated H₂ SO₄. Afterstirring for 2 days at 23°, the solution was poured onto ice andextracted into EtOAc. The organic phase was washed with saturated NaCland then with saturated NaHCO₃ until basic. The organic phase, whichstill contained the product, was washed with saturated NaCl, 12% HCl,and saturated NaCl. The organic phase was dried over MgSO₄, andevaporated under reduced pressure to a glass, 7.48 g. The structure wasconfirmed by NMR and mass spectroscopy. The material was of suitablepurity for use in the following step.

INTERMEDIATES FOR EXAMPLES 9 AND 10 DNMA-NHCH(OC₂ H₅)CO₂ C₂ H₅

DNMA-NHCH(OH)CO₂ H (3.0 g, 7.25 mmole) was dissolved in 100 ml absoluteEtOH. Concentrated H₂ SO₄ (1 ml) was added, and the mixture was stirredat 23° for 2 days. The mixture was evaporated under reduced pressure toan oil containing suspended solids. The mixture was taken into Et₂ O,washed with saturated NaCl and filtered to remove a minor amount ofinsoluble solids. The organic phase of the filtrate was washed withsaturated NaHCO₃ solution, saturated NaCl, 1N citric acid, and saturatedNaCl. The solution was dried over MgSO₄, and evaporated to a white foam,2.94 g. The foam was chromatographed on silica gel, eluting withEtOAc/CHCl₃ (1/1), giving the product as a white foam, 1.39 g, 40.9%yield. The structure was confirmed by NMR and mass spectroscopy. Thematerial was of suitable purity for use in the following step.

DNMA-NHCH(OC₂ H₅)CO₂ H

DNMA-NHCH(OC₂ H₅)CO₂ C₂ H₅ (1.27 g, 2.70 mmole) was dissolved in 45 mldioxane. 1N NaOH (5.7 ml) was added, and the mixture was stirred for 1.5hours. 1N HCl (6 ml) was added and the mixture was evaporated in vacuoto an oily residue.

The oil was taken up into EtOAc and washed with 1N citric acid,saturated NaCl, and dried over MgSO₄. The solution was evaporated to awhite foam, 1.37 g. The structure was confirmed by NMR and massspectroscopy. The product was of suitable purity for use in thefollowing step.

INTERMEDIATE FOR EXAMPLE 11 DNMA-NHCH(NH(CH₂)₃ NH₂)CO-STA-NHCH₂CH(CH₃)CH₂ CH₃

DNMA-NHCH(SCH₃)CO-STA-NHCH₂ CH(CH₃)CH₂ CH₃ (3.08 g, 4.6 mmole) wasdissolved in 150 ml dry THF and 1,3-diaminopropane (8 ml) was added.HgCl₂ (1.88 g, 6.9 mmole) was added and the suspension was stirred at23° overnight. The mixture was filtered and the filtrate evaporated toan oil. The oil was dissolved in EtOAc and washed with 12% HCl,saturated NaCl, 1N NaOH and saturated NaCl. The organic phase was driedover MgSO₄ and evaporated under reduced pressure to a white solid, 3.13g. The structure was confirmed by NMR and mass spectroscopy. Thematerial was used in the following step without further purification.

INTERMEDIATES FOR EXAMPLE 14 DNMA-NHCH(SPh)CO₂ H

DNMA-NACH(OH)CO₂ H (3.0 g, 7.25 mmole) was dissolved in 90 ml glacialHOAc and cooled to 0°. Thiophenol (1.5 ml, 14.5 mmole) was added,followed by concentrated H₂ SO₄ (10 ml). After stirring at 23° for fivedays, the HOAc was removed under reduced pressure and the residue wassuspended in EtOAc. The mixture was washed with saturated NaCl and thenwith saturated NaHCO₃ until washes were basic. The organic phase, whichstill contained the product, was washed with saturated NaCl, 1N citricacid, and saturated NaCl. The solution was dried over MgSO₄ andevaporated under reduced pressure to a white foam, 3.97 g. The foam wasdissolved in Et₂ O and precipitated by addition of hexane giving a gum.The supernatant liquid was decanted, and the residual gum wasredissolved in Et₂ O. By addition of a small amount of hexane, an orangeoil precipitated as an impurity. The solution was decanted from the oiland evaporated under reduced pressure giving the product as white foam,1.86 g. The structure was confirmed by NMR and mass spectroscopy. Thematerial was used without further purification in the following step.

INTERMEDIATES FOR EXAMPLES 18 AND 19 ##STR38##

A solution of 66 g (0.4 mole) of PHE in 120 ml of 3.33 N NaOH wastreated dropwise over 30 minutes with a solution of 37.1 g (0.2 mole) ofmorpholinosulfamyl chloride (prepared according to the method of R.Wegler and K. Bodenbennen, Ann. 624, 25 (1959)) in 80 ml of THF. Thesolution was stirred at room temperature for 6 hours, then acidified topH 2 with concentrated HCl. The mixture was extracted with EtOAc. TheEtOAc phase was washed with 1N HCl, dried over MgSO₄, and evaporated toa solid. Recrystallization from H₂ O gave 27 g of the pure product, m.p.157-158°. ##STR39## (10 g, 31.8 mmole) was dissolved in 250 ml CH₂ Cl₂/THF (1/1) and cooled to -50°. Carbonyldiimadazole (5.4 g, 33.4 mmole)was added and the mixture was warmed to -15° for 3 hours. The mixturewas purged with anhydrous NH₃ gas for 1 hour, followed by warming to 20°for 2 hours. The mixture was evaporated to a gelatinous mass andpartitioned between Et₂ O and H₂ O. The solid suspended in the Et₂ Ophase was collected and washed with H₂ O and Et₂ O. There was obtained6.0 g of the product as a crystalline solid. The structure was confirmedby NMR and mass spectroscopy. The material was used in the followingstep without further purification. ##STR40## (5.9 g, 18.8 mmole) wasdissolved in 300 ml of acetone. Glyoxylic acid.H₂ O (3.64 g, 39.5 mmole)was added and the mixture was heated at reflux for two days. The solventwas removed in vacuo, the residue dissolved in EtOAc, and washed with aminimal amount of saturated NaCl. The organic phase was then washedtwice with saturated NaHCO₃ solution, the washes combined, and acidifiedwith concentrated HCl to a Congo red end point. The mixture wasconcentrated in vacuo to a solid, mixed with EtOAc, and filtered. Thefiltrate was washed with saturated NaCl, dried over HgSO₄ and evaporatedunder reduced pressure to a white foam, 5.27 g. The structure wasconfirmed by NMR and mass spectroscopy. The material was used in thefollowing step without further purification. ##STR41## (5.11 g, 13.2mmole) was dissolved in 100 ml absolute EtOH. 1.0 ml concentrated H₂ SO₄was added, and the mixture stirred at 23° for five days. The mixture wasevaporated under reduced pressure to an oil, the oil dissolved in EtOAc,and washed with saturated NaCl, saturated NaHCO₃, and saturated NaCl.The EtOAc solution was dried over MgSO₄, and evaporated under reducedpressure to a glass, 5.54 g. The material was chromatographed on silicagel, eluting with EtOAc/CHCl₃ (1/1), to give 4.22 g of the product as aglass. The structure was confirmed by NMR and mass spectroscopy. Thematerial was used in the following step without further purification.##STR42##

A solution of 4.11 g ##STR43## in 50 ml EtOH was treated with 10 ml or1N NaOH and stirred for 0.5 hours. An additional 10 ml of 1N NaOH wasthen added and the stirring continued for an additional 0.5 hours. Themixture was acidified with 13 ml of 1N HCl and the solvent removed underreduced pressure. The residue was taken up in EtOAc, washed with 1N HCl,then saturated NaCl. Drying and removal of the solvent under reducedpressure gave 3.77 g of the product as a white foam. The structure wasconfirmed by NMR and mass spectroscopy. The material was used in thenext step without further purification.

COMMON INTERMEDIATES DNMA-Cl

Di-(1-naphthylmethyl)acetic acid (46.8 g, 0.137 mole) was dissolved withwarming to 30° in 120 ml thionyl chloride. After stirring overnight, thesolvent was removed under reduced pressure until the residual oil beganto crystallize. 400 ml Et₂ O was then added, giving a suspension, towhich was added 500 ml hexane. The mixture was concentrated underreduced pressure, cooled and filtered. The solid was washed with hexaneand dried giving the product as a white solid, 42.94 g. The structurewas confirmed by NMR and mass spectroscopy. The material was of suitablepurity for use in the following reaction.

DNMA-NH₂

150 ml THF was cooled to -60° and saturated with anhydrous NH₃ gas.While continuing the NH₃ purge, a solution of DNMA-Cl (16.0 g, 0.045mole) in 125 ml THF was added over 15 minutes. A solid formed, and theNH purge was discontinued. After warming to 23° overnight, the mixturewas stripped to a solid and taken up in 350 ml EtOAc. The EtOAc waswashed with 1N citric acid, saturated NaCl solution, saturated NaHCO₃solution, and saturated NaCl solution. The organic phase was dried overMgSO₄ and stripped under reduced pressure to a solid. The solid wasrecrystallized from EtOAc/Et₂ O, filtered, and dried, giving the productas a white solid, 13.4 g, 88% yield. The structure was confirmed by NMRand mass spectroscopy. The material was of sufficient puritry for use inthe following step.

DNMA-NHCH(OH)CO₂ H

DNMA-NH₂ (12.9 g, 0.038 mole) and glyoxylic acid hydrate (4.34 g, 0.047mole) were dissolved in 250 ml acetone and heated to reflux for 18hours. The mixture was evaporated to an oil, the oil dissolved in Et₂ Oand washed with saturated NaCl. Upon washing with saturated NaHCO₃, asolid precipitated which was filtered, triturated with EtOAc andresuspended in EtOAc. An equal volume of water was added, and pH wasadjusted to a Congo red end point by the addition of citric acid.Solution occurred and the phases were separated. The organic phase waswashed with saturated NaCl, dried over MgSO₄, and evaporated to a foam,12.7 g, 80% yield. The structure was confirmed by NMR and massspectroscopy. The material was of suitable purity for use in thefollowing step.

BOC-STA-NHCH₂ CH(CH₃)CH₂ CH₃

BOC-STA (27.53 g, 0.1 mole, US Patent 4,397,786) and HOBT.H₂ O (14.2 g,0.105 mole) were dissolved in 40 ml DMF. 300 ml CH₂ Cl₂ was added, andthe mixture was cooled to 0°. A solution of DCC (21.66 g, 0.105 mole) in50 ml CH₂ Cl₂ was added, followed by S-2-methylbutylamine (12 ml, 0.1mole). After stirring at 0° for 2 hours, the mixture was allowed to warmto 25° over 1.5 hours. The mixture was filtered, and the solvent wasremoved in vacuo. The residue was dissolved in EtOAc, which was washedwith 1N citric acid, brine, saturated NaHCO₃ solution, and brine. Theorganic phase was dried over MgSO₄, filtered, and stripped to a gum,36.90 g. The gum was dissolved in Et₂ O and treated with charcoal toremove colored impurities. The suspension was filtered and stripped to agum, 35.2 g, which was suitable for use in the following procedurewithout further purification.

STA-NHCH₂ CH(CH₃)CH₂ CH₃

A solution of 38.0 g, (0.11 mole) of BOC-STA-NHCH₂ CH(CH₃)CH₂ CH₃ in 250ml CH₂ Cl₂ was treated with HCl gas every one-half hour over a 3 hourperiod. The solvent was removed under reduced pressure and the residuetaken up in 30 ml H₂ O and 110 ml of 1N HCl. The solution was washedtwice with Et₂ O, the pH brought to 13 with 1N NaOH, and the solutionextracted twice with Et₂ O. The Et₂ O was washed with saturated NaCl,dried, and the solvent removed under reduced pressure giving 22.3 g ofthe product as an oil which solidified on standing. The material wassufficiently pure for use in subsequent reactions. ##STR44##

A solution of 17.7 g (0.056 mole) of BOC-STA and 7.59 g (0.056 mole) ofHOBT in 250 ml of DMF was cooled in ice and treated with a solution of11.7 g (0.056 mole) of DCC in 20 ml of DMF. After stirring for 5minutes, the solution was treated with 7.6mml (0.056 mole) of4-(2-aminoethyl)morpholine. The solution was stirred for 0.5 hours at0°, then at room temperature overnight. The dicyclohexylurea wasfiltered off and the solvent removed under high vacuum. The residue wastaken up in EtOAc, washed with saturated NaHCO₃, H₂ O, then saturatedNaCl, and then dried over MgSO₄. Removal of the solvent under reducedpressure gave the crude product which was purified by chromatography onsilica gel, eluting with CHCl₃ /MeOH (95/5). There was obtained 24 g ofpure product.

Similarly, substituting BOC-CHSTA (DE-3,610,593), BOC-ACYS(Z)(EP-198,271), BOC-DFKCYS (EP-222, 523), or BOC-DFKSTA (GB-2,171,103) forBOC-CYSTA in the above preparation gives the corresponding amides,##STR45##

A solution of 24 g (0.056 mole) of ##STR46## in 300 ml of CH₂ Cl₂ wastreated with HCl gas for 5 minutes. A gum formed which was redissolvedby the addition of 100 ml of MeOH. HCl gas was bubbled in for anadditional 10 minutes, and the solution allowed to stir at roomtemperature for 3 hours. The solvent was removed under reduced pressureand the residue taken up in CH₂ Cl₂. This was treated with CH₂ Cl₂ thathad been saturated with NH₃ gas at 0°. The NH₄ Cl was filtered off andthe filtrated evaporated. There was obtained 15.2 g of an oil whichsolidified on standing. The structure was confirmed by NMR spectroscopy.

Substitution of ##STR47## in the above preparation gives thecorresponding deviatives, ##STR48##

These may be converted to certain peptides of the present invention byfollowing the procedures outlined in Examples 18 and 19. In the case ofthe ##STR49## derivative, a final removal of the Z-group from theprecursor peptide gives certain other peptides of the present invention.

We claim:
 1. A peptide of the formula

    ACYL--X--Y--W--U--V                                        (I)

or a pharmaceutically acceptable acid addition salt thereof, whereinacyl is BOC, IVA, DNMA, BBSP, ##STR50## wherein R and R' are eachindependently hydrogen or straight or branched chain lower alkyl,##STR51## is a saturated ring containing 1 to 5 carbon atoms wherein Qis CH₂, O, or NR; X is absent, PHE, NAPHTHYLALA, or O-MeTYR, with theproviso that when ACYL is DNMA, X is absent; Y is ##STR52## wherein R₁is lower alkyl, lower alkenyl, lower alkynyl, aryl, (CH₂)_(n) --NHR₂,wherein n is an integer of from 2 to 4, and R₂ is ##STR53## wherein R₇is R₁, ##STR54## H, wherein R₈ is lower alkyl and R₉ is alkyl oraralkyl; W is STA, CYSTA, DFKSTA, DFKCYS, U is absent, and V is --NHCH₂Ph, ##STR55## ##STR56## ##STR57##
 2. A compound according to claim 1wherein in Y, R₁ and R₇ are methyl, ethyl, isopropyl, propyl, allyl,propargyl, phenyl, or (CH₂)_(n) --NHR₂, wherein n is an integer of from2 to 4 and R₂ is ##STR58##
 3. A compound according to claim 1 wherein inY, R₇ is H, ##STR59## where R₆ is methyl,.
 4. A compound according toclaim 1 wherein ACYL is BOC, IVA, or DNMA.
 5. A compound according toclaim 1 wherein V is -NHCH₂ Ph, ##STR60##
 6. A compound according toclaim 1 wherein W is STA, CYSTA, DFKCYS or DFKSTA.
 7. A compoundselected from the group consisting of:DNMA-NHCH(OCH₃)CO-STA-NHCH₂CH(CH₃)CH₂ CH₃, DNMA-NHCH(SCH₃)CO-STA-NHCH₂ CH(CH₃)CH₂ CH₃,DNMA-NHCH(SOCH₃)CO-STA-NHCH₂ CH(CH₃)CH₂ CH₃,DNMA-NHCH(SCH(CH₃)₂)CO-STA-NHCH₂ CH(CH₃)CH₂ CH₃ (fast isomer).DNMA-NHCH(SCH(CH₃)₂)CO-STA-NHCH₂ CH(CH₃)CH₂ CH₃ (slow isomer),DNMA-NHCH(NH₂)CO-STA-NHCH₂ CH(CH₃)CH₂ CH₃ (fast isomer),DNMA-NHCH(NH₂)CO-STA-NHCH₂ CH(CH₃)CH₂ CH₃ (slow isomer), DNMA-NHCH(NHCO₂Et)CO-STA-NHCH₂ CH(CH₃)CH₂ CH₃, DNMA-NHCH(OC₂ H₅)CO-STA-NHCH₂ CH(CH₃)CH₂CH₃ (fast isomer), DNMA-NHCH(OC₂ H₅)CO-STA-NHCH₂ CH(CH₃)CH₂ CH₃ (slowisomer), DNMA-NHCH(NH(CH₂)₃ NHCSNHCH₃)CO-STA-NHCH₂ CH(CH₃)CH₂ CH₃,DNMA-NHCH(NHCOCH₃)CO-STA-NHCH₂ CH(CH₃)CH₂ CH₃,DNMA-NHCH(NHPh)CO-STA-NHCH₂ CH(CH₃)CH₂ CH₃, DNMA-NHCH(SPh)CO-STA-NHCH₂CH(CH₃)CH₂ CH₃, DNMA-NHCH(N(CH₃)₂)CO-STA-NHCH₂ CH(CH₃)CH₂ CH₃ (slowisomer), DNMA-NHCH(NHCH(CH₃)₂)CO-STA-NHCH₂ CH(CH₃)CH₂ CH₃ (slow isomer),DNMA-NHCH(NHC₂ H₅)CO-STA-NHCH₂ CH(CH₃)CH₂ CH₃ (slow isomer), ##STR61##8. A pharmaceutical composition comprising a renin-inhibitory effectiveamount of a compound as claimed in claim 1 together with apharmaceutically acceptable carrier.
 9. A method of treatingrenin-associated hypertension which comprises administering to a mammala pharmaceutical composition as claimed in claim
 1. 10. A pharmaceuticalcomposition comprising an amount effective for treating congestive heartfailure of a compound according to claim 1 together with apharmaceutically acceptable carrier.
 11. A method of treating congestiveheart failure which comprises administering to a mammal a pharmaceuticalcomposition as claimed in claim 1.